Compounds which affect mRNA stability and uses therefor

ABSTRACT

Compounds which induce degradation of mRNA which contains one or more mRNA instability sequences are provided for use as pharmaceuticals, e.g. for use in the prophylaxis or treatment of diseases and medical conditions in general having an etiology associated with the increased or prolonged stability of mRNAs which contain one or more mRNA instability sequences, and which on prolonged or inappropriate expression typically give rise to undesirable effects, e.g. cancer cell growth or an unwanted inflammatory response.

CROSS REFERENCE TO RELATED APPLICATION

This is the 371 national stage of International Application No.PCT/CA99/01234, filed Dec. 23, 1999, published in English on Jul. 6,2000.

This invention relates to biologically active compounds and to their usein the treatment and prophylaxis of disease. In particular the inventionrelates to compounds which affect the stability of mRNA which containone or more mRNA instability sequences.

Recently, it has become increasingly apparent that the regulation of RNAhalf-life plays a critical role in the tight control of gene expressionand that mRNA degradation is a highly controlled process. RNAinstability allows for rapid up- or down-regulation of mRNA transcriptlevels upon changes in transcription rates. A number of criticalcellular factors, e.g. transcription factors such as c-myc, or geneproducts which are involved in the host immune response such ascytokines, are required to be present only transiently to perform theirnormal functions. Transient stabilisation of the mRNAs which code forthese factors permits accumulation and translation of these messages toexpress the desired cellular factors when required; whereas, undernon-stabilised, normal conditions the rapid turnover rates of thesemRNAs effectively limit and “switch off” expression of the cellularfactors. However, abnormal regulation of mRNA stabilisation can lead tounwanted build up of cellular factors leading to undesirable celltransformation, e.g. tumour formation, or inappropriate and tissuedamaging inflammatory responses.

Although the mechanisms which control mRNA stability are far fromunderstood, sequence regions have been identified in a number of mRNAs,which appear to confer instability on the mRNAs which contain them.These sequence regions are referred to herein as “mRNA instabilitysequences”. For example, typical mRNA instability sequences are the AREs(AU rich elements), which are found in the 3′UTR (3′ untranslatedregion) of certain genes including a number of immediate early genes andgenes coding for inflammatory cytokines, e.g. IL-1β and TNFα.

Kastelic et al. (CYTOKINE, Vol. 8, No. 10, (October), 1996: pp751-761)have reported the finding that radicicol analog A, if added to THP-1cells activated by IFN-Y and LPS, not only inhibited the secretion ofIL-1β but also induced an extremely rapid degradation of IL-1β, IL-6 andTNF-α mRNA to undetectable levels in 5-8 h, and that this mRNAdegradation appears to be mediated through AU-rich regions present inthe 3′ untranslated regions of the RNAs which code for these cytokines.

Previously, novel Radicicol analogs (including radicicol analog A),processes for their preparation and their pharmaceutical use weredescribed in European patent application EP 0606044 A, together withknown compounds including radicicol, O-methyl radicicol, and the relatedcompound zearelenone and certain analogs of zearelenone. The radicicolanalogs and known compounds are described in EP 0606044 A to be usefulfor the treatment of disorders with an etiology associated with orcomprising excessive cytokine release, particularly IL-1β release, suchas rheumatoid arthritis, osteoarthritis, septic shock, psoriasis,atherosclerosis, inflammatory bowel disease, Crohn's disease and asthma.

We have now found that there are other compounds in addition toradicicol analog A which induce degradation of mRNAs and that suchcompounds may be used for treatment of diseases and medical conditionswhich involve increased or prolonged stability and expression of suchmRNAs. Moreover we have found that radicicol analog A may be usedgenerally to induce degradation of mRNAs besides IL-1β, IL-6 and TNF-αmRNAs.

Accordingly the present invention provides a compound which inducesdegradation of mRNA which contains one or more mRNA instabilitysequences for use as a pharmaceutical, provided the compound is notradicicol analog A.

In a further aspect the invention provides a method for the prophylaxisor treatment of a disease or medical condition having an etiologyassociated with the increased stability of mRNA which contains one ormore mRNA instability sequences, comprising administering to a human oranimal patient an effective amount of a compound which inducesdegradation of the mRNA, provided that the compound is not radicicolanalog A when the disease or medical condition is one with an etiologyassociated with or comprising excessive cytokine release, particularlyIL-1β release, such as rheumatoid arthritis, osteoarthritis, septicshock, psoriasis, atherosclerosis, inflammatory bowel disease, Crohn'sdisease and asthma.

In a yet further aspect the invention provides use of a compound whichinduces degradation of mRNA which contains one or more mRNA instabilitysequences, for the preparation of a medicament for use in the treatmentor prophylaxis of a disease or medical condition having an etiologyassociated with the increased stability of mRNA which contains one ormore mRNA instability sequences, provided that the compound is notradicicol analog A when the disease or medical condition is one with anetiology associated with or comprising excessive cytokine release,particularly IL-1β release, such as rheumatoid arhritis, osteoarthritis,septic shock, psoriasis, atherosclerosis, inflammatory bowel disease,Crohn's disease and asthma.

The invention also provides a method for inducing degradation of mRNA ina patient, which comprises administering an effective amount of acompound which induces mRNA degradation to the patient, wherein the mRNAcontains an mRNA instability sequence, provided that the compound is notradicicol analog A when the mRNA is mRNA coding for IL-1β, IL-6 orTNF-α.

Further the invention provides use of a compound which induces mRNAdegradation in the preparation of a medicament for use in inducingdegradation of mRNA which contains a mRNA degradation sequence in apatient, provided that the compound is not radicicol analog A when themRNA is mRNA coding for IL-1β, IL-6 or TNF-α.

The present invention further provides the use of a radicicol analog forpreparation of a medicament for treatment of a cancer and/or malignantdisease.

The present invention also provides a method for the prophylaxis ortreatment of a cancer and/or malignant disease comprising administeringto a patient an effective amount of a radicicol analog.

Any compound which induces degradation of mRNA which contains a mRNAinstability sequence is potentially of interest for use in the presentinvention. Compounds which induce degradation of mRNA which contains amRNA instability sequence are hereinafter referred to as Compounds foruse in the invention. Such compounds include radicicol analogs, inparticular radicicol analog A or radicicol; for instance, as describedin EP 0606044.

Our copending British patent application no. 9828709.7 describes areporter gene assay for the identification of compounds whichdestabilise mRNA. In this assay test compounds are contacted with a DNAexpression system which in the absence of the compound is capable ofexpressing a protein having a detectable signal, and wherein the mRNAwhich codes for the protein and which is transcribed from the expressionsystem comprises at least one copy of a mRNA instability sequence. Thedetectable signal is measured in the presence of the test compound andthe result obtained is compared with a control. Compounds whichdestabilise mRNAs induce degradation of the mRNA which codes for thedetectable signal leading to a decrease in the magnitude of thedetectable signal obtained in the reporter gene assay.

Preferred compounds for use in the present invention include compoundswhich may be identified as inducers of mRNA instability using thereporter gene assay as described above and as described in more detailin the above mentioned British patent application no. 9828709.7 and ashereinafter described in the Examples. Particular examples of compoundsfor use in the present invention include radicicol and the radicicolanalogs.

Radicicol, the compound of formula I

has been known for many years as a natural compound, e.g. as ametabolite of the microorganism Monosporium bonorden, and was describedinitially as having antibiotic properties (Delmotte, Nature 171, 344(1953)).

A particular class of radicicol analogs which includes Compounds for usein the invention are compounds of formula II

wherein

R₁ is H, OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—;

R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—;

R₃ is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—;

—a—b— is —CHR₇—CHR₈— or cis or tans —CR₇═CR₈,

wherein R₇ and R₈ are the same or different and are H, OH, C₁-C₄ loweralkoxy, or C₁-C₄ lower alkyl-COO—, or

—a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge;

c is >CH—OH, >C═O or >CH₂;

—de is —CHR₇—CHR₈— or cis or trans —CR₇═CR₈—,

wherein R₇ and R₈ are the same or different and are H, OH, C₁-C₄ loweralkoxy, or C₁-C₄ lower alkyl-COO—, and

—f—g— is —CH₂—CH₂—, cis or trans —CH═CH—, or —C(O)—CH₂—,

and pharmaceutically acceptable salts thereof andphysiologically-hydrolysable and -acceptable esters thereof.

The carbon atom marked with an asterisk (*) in formula II is anasymmetric carbon atom. The carbon atoms at a, b, c or d may also beasymmetric carbon atoms dependent upon the particular substituentspresent at these positions. Asymmetric carbon atoms at these positionsmay have the R- or S-configuration or the radicicol analog may compriseany mixture of the optical isomers thereof. Preferred isomers includethose specifically described hereinafter.

Halogen or halo as used herein refers to F, Cl, Br or I unless otherwiseindicated, preferably Cl.

A particular subset of the compounds of formula II are those in whichone of —a—b— or —d—e— is —CHR₇—CHR₈— and the other is cis- or trans-—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and c is >CH—OH or >C═O,and wherein R₁, R₂, R₃ and —f—g— are as defined above.

Particular significances for the variable substituents and moieties ofthe radicicol analogs of formula II are as follows:

Preferably R₁ and R₃ are the same or different and are H, —OH, MeO— orMe—COO—. Preferably R₂ is —OH, MeO— or Me—COO—. More preferably R₁ is Hor MeO; R₂ is MeO, and R₃ is OH or MeO.

Preferably —a—b— is cis- or trans- —CR₇′═CR₈′—, wherein R₇′ and R₈′ arethe same or different and are H, OH, MeO— or Me—COO—. More preferably—a—b— is cis- or especially trans- —CH═CH—.

Preferably -de- is —CHR₇′—CHR₈′—, wherein R₇′ and R₈′ are as definedabove. More preferably -de- is —CH₂—CH₂— or especially —CHOH—CHOH—,wherein the OH groups may be in free or protected form.

Most preferably —f—g— is trans- —CH═CH—.

Preferably the asymmetric carbon atoms of the compounds of the inventionall have the S-configuration.

Particular radicicol analogs of formula II for use in the invention areanalogs of formula II in which R₁ is H or methoxy, R₂ is methoxy, R₃ isOH, —a—b— is cis- or trans-—CH═CH—, c is CHOH or C═O, —d—e— is—CHOH—CHOH— and —f—g— is trans- —CH═CH—; in free form or base salt formor in the form of a physiologically-hydrolysable and -acceptable ester.

Particular radicicol analogs for use in the present invention includeradicicol and O-lower alkyl radicicols, i.e. the compounds of formula I′

where R is H or C₁-C₄ lower alkyl, e.g. methyl, and pharmaceuticallyacceptable salts thereof and physiologically-hydrolysable and-acceptable esters thereof.

For the purposes of the present description a radicicol analog is acompound having the characteristic bicyclic ring structure of radicicol,i.e. the structure of formula I″,

wherein the X groups are separately H or substituents, the 14-memberedlactam ring may additionally comprise one or more, e.g. two,ethylenically unsaturated bonds and at least one of the X substituentsof the lactam ring may comprise an oxy (═O), or (with an adjacent Xsubstituent) an epoxide substituent, and pharmaceutically acceptablesalts thereof and physiologically-hydrolysable and -acceptable estersthereof.

Radicicol analogs which comprise —OH substituents may also exist in theform of pharmaceutically acceptable esters, and the use of such isincluded within the scope of the invention. Pharmaceutically acceptableesters are preferably prodrug ester derivatives, such being convertibleby solvolysis or under physiological conditions to the free radicicolanalog. Preferred pharmaceutically acceptable prodrug esters of the arethose derived from a carboxylic acid, a carbonic acid monoester or acarbamic acid, advantageously esters derived from an optionallysubstituted lower alkanoic acid or an arylcarboxylic acid.

Radicicol analogs may also exist in the form of pharmaceuticallyacceptable salts, and the use of such is included within the scope ofthe invention. Pharmaceutically acceptable salts represent acid additionsalts with conventional acids, for example, mineral acids, e.g.,hydrochloric acid, sulfuric or phosphoric acid, or organic acids, forexample, aliphatic or aromatic carboxylic or sulfonic acids, e.g.,acetic, propionic, succinic, glycolic, lactic, malic, tartaric, citric,ascorbic, maleic, fumaric, hydroxymaleic, pyruvic, pamoic,methanesulfonic, toluenesulfonic, naphthalenesulfonic, sulfanilic orcyclohexylsulfamic acid; also amino acids, such as arginine and lysine.For compounds of the invention having acidic groups, for example, ianacidic —OH group, pharmaceutically acceptable salts also represent metalor ammonium salts, such as alkali metal or alkaline earth metal salts,e.g., sodium, potassium, magnesium or calcium, salts.

EP 0606044 A describes the isolation and characterisation of theradicicol analog of formula III,

hereinafter referred to as radicicol analog A, which was firstidentified as a natural product isolated from a strain of pycnidiaimperfect fungi (F/87-250904) deposited on Nov. 6, 1991 with the ARSPatent Culture Collection, US Dept. of Agriculture, Northern RegionalResearch Centre, Peoria, Ill., USA under the provisions of the BudapestTreaty as deposit NRRL 18919.

Radicicol analog A is a particularly preferred radicicol analog for usein the present invention. Radicicol analog A also serves as a valuablestarting material for synthesis of other radicicol analogs for use inthe present invention. Alternatively EP 0606044 A describes the de novosynthesis of radicicol analogs starting from readily available startingmaterials.

Novel Radicicol analogs, processes for their preparation and theirpharmaceutical use are described in European patent application EP0606044 A, together with known compounds including radicicol, O-methylradicicol, and the related compound zearelenone and certain analogs ofzearelenone. The radicicol analogs and known compounds are described inEP 0606044 A to be useful for the treatment of disorders with anaetiology associated with or comprising excessive cytokine release,particularly IL-1β release, such as rheumatoid arthritis,osteoarthritis, septic shock, psoriasis, atherosclerosis, inflammatorybowel disease, Crohn's disease and asthma.

The disclosure of EP 0606044 relating to the isolation of radicicolanalog A from the fungal strain F/87-250904, the synthesis ofsemi-synthetic radicicol analogs from radicicol analog A and the de novosynthesis of radicicol analogs, is specifically incorporated byreference in the teaching of the present application.

Surprisingly it has now been found that radicicol, radicicol analogs,zearelenone and zearelenone analogs (hereinafter collectively referredto as radicicol analogues), such as those described in EP 0606044 A, areuseful for treatment of certain forms of cancer and malignant diseases.

Particularly preferred radicicol analogs for use in the inventioninclude compounds of formula II in which —a—b— is trans- —CH═CH—, e.g.the compounds of formulae IV, V and VI

Particularly preferred radicicol analogs for use in the inventioninclude compounds of formula II in which —a—b— is trans- —CH═CH—, e.g.the compounds of formulae III, VII and VIII.

The present invention may be used in the prophylaxis or treatment ofdiseases and medical conditions in general having an etiology associatedwith the increased or prolonged stability of mRNAs which contain one ormore mRNA instability sequences, and which on prolonged or inappropriateexpression typically give rise to undesirable effects, e.g. cancer cellgrowth or an unwanted inflammatory response.

mRNA instability sequences have been identified in the UTRs, inparticular the 3′UTRs, of a large number of transiently expressed genesincluding genes for cytokines, chemokines, nuclear trascription factors,protooncogenes, immediate early genes, cell cycle controlling genes,oxygenases, and genes involved in and controlling of apoptosis. Thenatural RNA sequences which comprise the mRNA instability sequences arealternatively referred to as adenylate/uridylate (AU)-rich elements, orAREs. Transiently expressed genes which contain mRNA instabilitysequences include, for example, the genes coding for GM-CSF, c-fos,c-myc, c-jun, krox-20, nur-77, zij268, β-IFN, uPA, IL-1, IL-3, TNF-α,MCP1, synl, β₂-AR, E-selectin, VCAM-1, ICAM-1, P-glycoproteins (MDR),MRPs, Pγhl (pf mdr), COX II, metalloproteinases (MMPs), bcl-2 and MIP-2α.

The following publications include extensive discussion of mRNAinstability sequences and AREs, the sequences motifs which they containand (minimum) sequence requirements for mRNA destabilisation, as well asidentifying a number of mRNA instability sequences and the genes whichcontain them:

Shaw & Kamen, Cell, Vol. 46, 659-667, Aug. 29, 1986 (GM-CSF);

Shyu et al., Genes & Development, 5:221-231 (1991) (c-fos);

Sachs, Cell, Vol. 74, 413-421, Aug. 13, 1993 (Review. “Messenger RNADegradation in Eukaryotes”);

Chen et al., Mol. Cell. Biol., January 1994, p416-426 (cfos);

Akashi et al., Blood, Vol. 83, No. 11, (Jun. 1), 1994: pp 3182-3187(GM-CSF etc.);

Nanbu et al., Mol. Cell. Biol., July 1994, p. 4920-4920 (Upa);

Stoecklin et al., J. Biol. Chem., Vol. 269, No. 46, Nov.18, 1994, pp28591-28597 (IL-3);

Lagnado et al., Mol. Cell. Biol., December 1994, p. 7984-7995 (general);

Zhang et al., Mol. Cell. Biol., April 1995, p. 2231-2244 (yeast);

Zubiaga et al., Mol. Cell. Biol., April 1995, p. 2219-2230 (general);

Winstall et al., Mol. Cell. Biol., July 1995, p. 3796-3804 (c-fos,GM-CSF);

Chen et al., Mol. Cell. Biol., October 1995, p. 5777-5788 (c-fos,GM-CSF);

Chen et al., TIBS Nov. 20, 1995, 465-470 (review);

Levy et al., J. Biol. Chem., Vol. 271, No. %, Feb. 2, 1996, pp.2746-2753 (VEGF);

Kastelic et al., Cytokine, Vol. 8, No. 10 (October), 1996: pp751-761;

Crawford et al., J. Biol. Chem., Vol. 272, No. 34, Aug.22, 1997, pp.21120-21127 (TNFα);

Xu et al., Mol. Cell. Biol., August 1997, Vol. 18, No. 8, p. 4611-4621(general);

Danner et al., J. Biol. Chem., Vol.273, No. 6, Feb.6, 1998, pp.3223-3229 (human β₂-Adrenergic Receptor);

Lewis et al., J. Biol. Chem., Vol. 273, No. 22, May 29 1998, pp.13781-13786 (TNF-α).

As described in the above publications mRNA instability sequences oftencontain one or more copies of sequence motifs, e.g. selected from:AUUUA, UAUUUAU, UUAUUUA(U/A)(U/A), and AUUUAUUUA. Such sequence motifsare typically in genes between the stop codon and the poly A signal andmay associated with appropriate flanking sequences and may interact incombination with other sequences, e.g. present in the 5′ UTR and e.g.with instability motifs present in the coding region.

The present invention may be used in connection with diseases andmedical conditions associated with any of the genes mentioned above ordescribed in the listed publications, which comprise mRNA instabilitysequences.

Examples of diseases and medical conditions which may be treated orprevented by use of the present invention include: cancers e.g. of thecolon, breast, lung etc., acute and chronic inflammation, autoimmunediseases, respiratory diseases, infectious diseases and transplantrejection.

The compounds for use in the invention have valuable pharmacologicalproperties. In particular compounds for use in the invention havevaluable properties as inducers of degradation of mRNAs which containmRNA instability sequences. The activity of compounds for use in theinvention as inducers of mRNA degradation may be demonstrated by meansof a reporter gene assay as hereinafter described in the Examples, or asdescribed in more detail in our copending British patent application no.9828709.7.

In view of their activity as inducers of degradation of mRNAs whichcontain mRNA instability sequences, the radicicol analogues are usefulfor the prophylaxis and treatment of cancers and malignant diseaseswhich involve inappropriate build-up and expression of mRNAs, whichcontain mRNA instability sequences, and which code for proteins involvedin the initiation, progression or persistence of cancer or malignantdisease. Examples of cancer related genes, with mRNAs which contain mRNAdestabilising sequences, include various oncogenes and transcriptionfactors, e.g. c-myc, c-fos, Spl, bcl-2 and similar genes. Theinappropriate or prolonged expression of such oncogenes is implicated inthe initiation of certain forms of cancer, such as colon cancer, breastcancer, lung cancer etc.. Further examples of cancer related genes, withmRNAs which contain mRNA instability sequences are genes formetalloproteinase enzymes, e.g. MMP-1, MMP-2, collagenases etc.,involved in tissue remodelling required for tumour growth and metastasisinvasion; cell cycle related genes such as p45/SKIP2 etc. and multidrugresistance genes, e.g. mdr-1, MRPs, etc. involved in the intrinsic oracquired multidrug resistance of some cancer cells.

Treatment with radicicol analogs advantageously leads to degradation ofthe m1RNAs of such genes, resulting in the down-regulation or “switchingoff” of gene expression. Thus for example, radicicol analogs may be usefor treatment and prevention of oncogene mediated cancers and malignantdiseases, to treat or prevent tumour growth and metastasis invasion ingeneral, and to prevent or reverse multidrug resistance and therebyfacilitate cancer and tumour treatment with conventional, e.g.cytotoxic, anti-cancer agents.

Radicicol analogs may be tested for their activity as anticancer agentsin cell or in vivo assays substantially as described below or invariants of such assays using appropriate cell lines and conditions.

Radicicol analogs exhibit, for example, inhibition of the cell growth ofEGF-dependent cell lines, for example the epidermoid BALB/c mousekeratinocyte cell line (see Weissmann, B. A., and Aaronson, S. A., Cell32 599 (1983)) or the A431 cell line, which are recognised usefulstandard sources of EGF-dependent epithelial cells (see Carpenter, G.,and Zendegni, J. Anal. Biochem. 153, 279-282 (1985)). In a known testmethod (see Meyer et al., Int. J. Cancer 43, 851 (1989)), the inhibitoryactivity of radicicol analogs is determined, briefly, as follows:BALB/MK cells (10 000/microtitre plate well) are transferred to 96 wellmicrotitre plates. The test compounds (dissolved in DMSO) are added in aseries of concentrations (dilution series) in such a manner that thefinal concentration of DMSO is not greater than 1% (v/v). After theaddition, the plates are incubated for three days during which thecontrol cultures without test compound are able to undergo at leastthree cell-division cycles. The growth of the MK cells is measured bymeans of methylene blue staining: after the incubation the cells arefixed with glutaraldehyde, washed with water and stained with 0.05%methylene blue. After a washing step the stain is eluted with 3% HCl andthe optical density per well of the microtitre plate is measured using aTitertek multiskan at 665 nm. IC₅₀ values are determined by acomputer-aided system using the formula:

IC₅₀=[(OD _(test) −OD _(start))/(OD _(control) −OD _(start))]×100.

The IC₅₀ value in those experiments is given as that concentration ofthe test compound in question that results in a cell count that is 50%lower than that obtained using the control without inhibitor. Theradicicol analogs exhibit inhibitory activity in the micromolar range,for example an IC₅₀ of approximately from 0.1 to 10 mM, especially from0.4 to 4 mM.

The radicicol analogs exhibit inhibition of the growth of tumour cellsalso in vivo, as shown, for example, by the test described below: thetest is based on inhibition of the growth of the human epidermoidcarcinoma A43 1 (ATCC No. CRL 1555; American Type Culture Collection,Rockville, Md., USA; see Santon, J. B., et al., Cancer Research 464701-4705 (1986) and Ozawa, S., et aL, Int. J. Cancer 40, 706-710(1987)), which is transplanted into female BALB/c nude mice(Bomholtgard, Denmark). That carcinoma exhibits a growth that correlateswith the extent of the expression of EGF-receptor. In the experiment,tumours having a volume of approximately 1 cm³ cultured in vivo aresurgically removed from experimental animals under sterile conditions.The tumours are comminuted and suspended in 10 volumes (w/v) ofphosphate-buffered saline. The suspension is injected s.c. (0.2 ml/mousein phosphate-buffered saline) into the left flank of the animals.Alternatively, 1×10⁶ cells from an in vitro culture can be injected in0.2 ml of phosphate-buffered saline. Treatment with test compounds isstarted 5 or 7 days after the transplant, when the tumours have reacheda diameter of 4-5 mm. The test compound in question is administered (indifferent doses for different animal groups) once a day for 15successive days. The tumour growth is determined by measuring thediameter of the tumours along three axes that are perpendicular to eachother. The tumour volumes are calculated using the known formulap×L×D²/6 (see Evans, B. D., et al., Brit. J. Cancer 45, 466-468 (1982)).The results are given as treatment/control percentages (TIC×100=T/C%).At a dose of from 3 to 50 mg/kg active ingredient, distinct inhibitionof the tumour growth is found, for example T/C% values of less than 10,which indicates strong inhibition of tumour growth.

The radicicol analogs for use in the invention can be used both aloneand in combination with other pharmacologically active compounds, forexample together with inhibitors of the enzymes of polyamine synthesis,inhibitors of protein kinase C, inhibitors of other tyrosine kinases,cytokines, negative growth regulators, for example TGF-β or IFN-β,aromatase inhibitors, antioestrogens and/or cytostatic agents.

Characteristically when the radicicol analogs are use to prevent orreverse multidrug resistance of tumour and other malignant cells, theyare used in combination with cytostatic or cytotoxic agents. A suitablecell-based assay for assessing utility in restoring sensitivity ofcancer cells to anti-neoplastic/cytotoxic, drug substances in vitro isas follows.

Cancer cell lines (CCL), e.g. from human small cell carcinoma of thelung, resistant to one or more cancer therapeutic drug substances (CTDS)selected from the group comprising Daunorubicin (DR); Vincristine (VC);Adriamycin (AM); Etoposide (ET); Tenoposide (TE); Colchicine (CC); andTaxol are developed in accordance with the methods described byTwentyman et al., Br. J. Cancer, 54, 253 (1986).

Sensitivity of resistant sub-lines (CCL-R) is compared with parentalsensitive lines (CCL-S)by assaying inhibition of cell growth duringcontinuous CTDS exposure, e.g. in the case of a DR-resistant line(CCL-DRR)by comparing growth of CCL-DRS and CCL-DRR lines in thepresence of DR contained in the growth medium ab initio. For thepurpose, cell proliferation is measured by cell counting using anelectronic cell counter, counting being effected close to thetermination of the exponential growth phase. CCL-R lines are selectedfor which the IC₅₀ (drug concentration, e.g. DR concentration, requiredto reduce final cell number to 20% of that for non-CTDS (e.g. DR)treated controls is >80×, preferably >100×, greater than that of theparental CCL-S lines.

Sensitivity of selected CCL-R lines to CTDS (e.g. DR) in the presence orabsence of test radicicol analog is then performed, employing cellcounting as a measure of proliferation as described above. For thispurpose cells are cultured ab initio in the presence of varyingconcentrations of both CTDS and test radicicol analog. For screening,concentrations of the latter are chosen which do not themselves cause asignificant reduction in proliferation. Appropriate concentrations areestablished by culturing CCL-S and CCL-R in the presence of varyingconcentrations of radicicol analog in the absence of CTDS. Radicicolanalogs are routinely tested at concentrations of from 0.01 to 50, inparticular 0.1 to 10 μg/ml, e.g. at concentrations of 0.01, 0.02, 0.05,0.1, 0.2, 0.5, 1.0, 2.0, 5.0, 10.0 and 50 μg/ml. The ratio of CTDS (e.g.DR) required to inhibit cell proliferation by 50% in the absence of testradicicol analog (IC₅₀−CS) compared with that obtained in the presenceof test radicicol analog (IC₅+CS) is taken as a measure of increasedsensitivity of the CCL-R line to CTDS which has been induced by theradicicol analog. Stability of the CCL-R line used is ensured by crosschecking its sensitivity to CTDS with that previously established.

Additional procedures for assessing utility in restoring sensitivity ofcancer cells to anti-neoplastic/cytotoxic, drug substances, including invivo procedures are described in EP 0296122 B, the relevant disclosuresof which are incorporated by reference in the teaching of the presentapplication.

Compounds for use in the Invention can be used both alone and incombination with other pharmacologically active compounds, for examplein cancer treatment the compounds may be used together with inhibitorsof the enzymes of polyamine synthesis, inhibitors of protein kinase C,inhibitors of other tyrosine kinases, cytokines, negative growthregulators, for example TGF-β or IFN-β, aromatase inhibitors,antioestrogens and/or cytostatic agents.

Suitable pharmaceutical compositions comprising Compounds for use in theinvention as active ingredient and that can be used especially in thetreatment of the diseases mentioned above include compositions forenteral, such as nasal, buccal, rectal or especially oral,administration and parenteral, such as intravenous, intramuscular orsubcutaneous, administration to warm-blooded animals, especially humanbeings. The compositions comprise the active ingredient on its own orpreferably together with a pharmaceutically acceptable carrier. Thedosage of the active ingredient depends on the disease to be treated,and on species, age, weight and individual condition, individualpharmacokinetic conditions, and the mode of administration.

The pharmaceutical compositions may comprise from approximately 1% toapproximately 95% active ingredient, forms of administration in singledose form preferably comprising from approximately 20% to approximately90% active ingredient and forms of administration that are not in singledose form preferably comprising from approximately 5% to approximately20% active ingredient. Unit dose forms are, for example, dragdes,tablets, ampoules, vials, suppositories or capsules. Other forms ofadministration are, for example, ointments, creams, pastes, foams,tinctures, lipsticks, drops, sprays, dispersions, etc. Examples arecapsules comprising from approximately 0.05 g to approximately 1.0 g ofthe active ingredient.

The pharmaceutical compositions are prepared in a manner known per se,for example by means of conventional mixing, granulating, confectioning,dissolving or lyophilising procedures.

Solutions of the active ingredient, and also suspensions or dispersions,especially isotonic aqueous solutions, dispersions or suspensions, arepreferably used, it being possible, for example in the case oflyophilised compositions that contain the active ingredient alone ortogether with a carrier, for example mannitol, for such solutions,suspensions or dispersions to be made up prior to use. Thepharmaceutical compositions may be sterilised and/or may compriseexcipients, for example preservatives, stabilisers, wetting agentsand/or emulsifiers, solubilisers, salts for regulating the osmoticpressure and/or buffers, and are prepared in a manner known per se, forexample by means of conventional dissolving or lyophilising procedures.The said solutions or suspensions may comprise viscosity-increasingsubstances, such as sodium carboxymethyl-cellulose, dextran,polyvinylpyrrolidone or gelatin.

Suspensions in oil comprise as the oil component the vegetable,synthetic or semi-synthetic oils customary for injection purposes. Theremay be mentioned as such especially liquid fatty acid esters thatcontain as acid component a long-chained fatty acid having from 8 to 22,especially from 12 to 22, carbon atoms, for example lauric acid,tridecylic acid, myristic acid, pentadecylic acid, palmitic acid,margaric acid, stearic acid, arachidic acid, behenic acid, orcorresponding unsaturated acids, for example oleic acid, elaidic acid,erucic acid, brassidic acid or linoleic acid, if desired with theaddition of antioxidants, for example vitamin E, β-carotene or3,5di-tert-butyl4-hydroxytoluene. The alcohol component of those fattyacid esters has a maximum of 6 carbon atoms and is a mono- orpoly-hydric, for example a mono-, di- or tri-hydric, alcohol, forexample methanol, ethanol, propanol, butanol or pentanol or the isomersthereof, but especially glycol and glycerol. The following examples offatty acid esters are therefore to be mentioned: ethyl oleate, isopropylmyristate, isopropyl palmitate, “Labrafil M 2375” (polyoxyethyleneglycerol trioleate, Gattefossd, Paris), “Labrafil M 1944 CS”(unsaturated polyglycolised glycerides prepared by alcoholysis ofapricot kernel oil and consisting of glycerides and polyethylene glycolester, Gattefossé, France), “Labrasol” (saturated polyglycolisedglycerides prepared by alcoholysis of TCM and consisting of glyceridesand polyethylene glycol ester, Gattefosse, France) and/or “Miglyol 812”(triglyceride of saturated fatty acids with a chain length of C₈ to C₁₂,Hüls AG, Germany), but especially vegetable oils, such as cottonseedoil, almond oil, olive oil, castor oil, sesame oil, soybean oil and moreespecially groundnut oil.

The injection compositions are prepared in customary manner understerile conditions; the same applies also to introducing thecompositions into, for example, ampoules or vials and to sealing thecontainers.

Pharmaceutical compositions for oral administration can be obtained, forexample, by combining the active ingredient with one or more solidcarriers, if desired granulating a resulting mixture, and processing themixture or granules, if desired, and if necessary by the addition ofadditional excipients, to form tablets or dragée cores.

Suitable carriers are especially fillers, such as sugars, for examplelactose, saccharose, mannitol or sorbitol, cellulose preparations and/orcalcium phosphates, for example tricalcium phosphate or calcium hydrogenphosphate, and also binders, such as starches, for example corn, wheat,rice or potato starch, methylcellulose, hydroxypropylmethylcellulose,sodium carboxymethylcellulose and/or polyvinylpyrrolidone, and/or, ifdesired, disintegrators, such as the above-mentioned starches, alsocarboxymethyl starch, crosslinked polyvinylpyrrolidone, or alginic acidor a salt thereof, such as sodium alginate. Additional excipients areespecially flow conditioners and lubricants, for example silicic acid,talc, stearic acid or salts thereof, such as magnesium or calciumstearate, and/or polyethylene glycol, or derivatives thereof.

Dragée cores can be provided with suitable, optionally enteric,coatings, there being used inter alia concentrated sugar solutions whichmay contain gum arabic, talc, polyvinylpyrrolidone, polyethylene glycoland/or titanium dioxide, or coating solutions in suitable organicsolvents or solvent mixtures, or, for the production of entericcoatings, solutions of suitable cellulose preparations, such asacetylcellulose phthalate or hydroxypropylmethylcellulose phthalate.Colourings or pigments may be added to the tablets or dragee coatings,for example for identification purposes or to indicate different dosesof active ingredient.

Orally administrable pharmaceutical compositions also include dry-filledcapsules consisting of gelatin, and also soft, sealed capsulesconsisting of gelatin and a plasticiser, such as glycerol or sorbitol.The dry-filled capsules may contain the active ingredient in the form ofgranules, for example in admixture with fillers, such as corn starch,binders and/or glidants, such as talc or magnesium stearate, andoptionally stabilisers. In soft capsules, the active ingredient ispreferably dissolved or suspended in suitable liquid excipients, such asfatty oils, paraffin oil or liquid polyethylene glycols or fatty acidesters of ethylene or propylene glycol, to which stabilisers anddetergents, for example of the polyoxyethylene sorbitan fatty acid estertype, may also be added.

Other oral forms of administration are, for example, syrups prepared incustomary manner which comprise the active ingredient, for example, insuspended form and in a concentration of about 5% to 20%, preferablyabout 10%, or in a similar concentration that provides a suitable singledose, for example, when administered in measures of 5 or 10 ml. Alsosuitable are, for example, powdered or liquid concentrates for thepreparation of shakes, for example in milk. Such concentrates may alsobe packaged in single dose quantities.

Suitable rectally administrable pharmaceutical compositions are, forexample, suppositories that consist of a combination of the activeingredient and a suppository base. Suitable suppository bases are, forexample, natural or synthetic triglycerides, paraffin hydrocarbons,polyethylene glycols or higher alkanols.

For parenteral administration there are suitable especially aqueoussolutions of an active ingredient in water-soluble form, for example inthe form of a water-soluble salt, or aqueous injection suspensions thatcontain viscosity-increasing substances, for example sodiumcarboxymethylcellulose, sorbitol and/or dextran, and, if desired,stabilisers. The active ingredient, optionally together with excipients,can also be in the form of a lyophilisate and can be made into asolution prior to parenteral administration by the addition of suitablesolvents.

The Compounds for use in the invention can be administered,prophylactically or therapeutically, as such or in the form ofpharmaceutical compositions, preferably in an amount effective againstthe said diseases, to a warm-blooded animal, for example a human being,requiring such treatment, the compounds being used especially in theform of pharmaceutical compositions. In such treatment an individual ofabout 70 kg body weight will be administered a daily dose of fromapproximately 0.1 g to approximately 5 g, preferably from approximately0.5 g to approximately 2 g, of a compound of formula II.

The following Examples serve to illustrate the invention and refer tothe accompanying Figures, in which

FIG. 1 which shows the 30 bp fragment used as a mRNA instabilitysequence in the porter gene assay of Example 1 (SEQ ID NOS: 1 and 2);

FIG. 2 which shows plasmid diagrams for pGL2_Neo30 and pGL2-Control; and

FIG. 3 shows graphs of luciferase activity from clones 53 (solid bars)and 63 (open bars) treated with various concentrations of radicicolanalog A (SDZ 216-732).

EXAMPLES Example 1 Reporter Gene Assay for Compounds Which DestabilisemRNA

A. Construction of pGL2 neo30

In order to obtain a vector for stable integration into THP-1 cells, aXhoI-SalI fragment of the neo resistant gene (expressing aminoglycoside3′ phosphotransferase) derived from pMCIneo (Stratagene) is subclonedinto the SalI site of pGL2Control (Promega). This resulting plasmid wascalled pGL2_Neo. A 30 bp fragment (containing three tandem AUUUA motifs,based on the IL-Iβ3′UTR sequence) obtained by annealing twocomplementary synthetic oligonucleotides (see FIG. 1) is subcloned intopGL2 Neo using the PflM1 restriction site. This results in theluciferase expression vector pGL2_Neo30 (FIG. 2). FIG. 1 shows theIL-1β3′UTR sequence containing three tandem AUUUA motifs used forligation into the PflMI site of pGL2_Neo.

B. Transfection and Selection of Stable Cell Lines

The resulting vectors pGL2_Neo30 and pGL2_Neo are transfected into THP-1cells by electroporation. 10⁷ cells/ml in 1.3 mM KH₂PO₄, 7.36 m.MNa₂HPO₄, 2.44 mM KCl, 124 mm NaCl, 5 mM glucose, 9.6 μM MgCl₂ and 16 μMCaCI₂ pH 7.2 are transfected with 20 μg of DNA in a Bio-Rad Gene Pulser(250V, 690 μF and indefinite resistance) using a 0.4 cm cuvette. Cellsare subsequently cultured in RPMI medium containing 10%FBS, 2mM L-Gln(L-glutamine), 50 μM 2mercaptoethanol and 600 μg/ml of G418 (geneticin).After transfection of pGL2_Neo30 and pGL2_Neo into THP-1 cells, stablecell lines are obtained by selection for G418 resistance and assayed forluciferase activity. One cell line of each transfection is chosen forfurther analysis; the pGL2_Neo30 cell line is referred to as clone No.63 and the pGL2_Neo cell line as clone No. 53. No endogenous luciferaseactivity could be detected in normal THP-1 cells.

The tissue culture and luciferase activity measurements are carried outas described below.

C. Tissue Culture

The transfected human monocytic leukemia cell lines, clones No. 53 and63 are grown in RPMI medium supplemented with 110 U/ml penicillin, 100μg/ml streptomycin, 2 mM L-Gln and 2 g/l NaHCO₃. Heat-treated FBS (5%)is added before use. The cells are grown to a density of 5×10⁵/ml andinduced to differentiate with 100 U/ml (final concentration) γIFN. Threehours later, 10 μl of LPS (5 μg/ml final concentration) is added. Thistime point is designated time 0. Compounds are added at various timesafter LPS addition as indicated.

D. Luciferase Activity Measurement

In order to adapt the system to the use of 96 well plates, cells aregrown in Packard flat bottom white polystyrene microplates (CatNo.6005180) in RPMI medium lacking phenol red (AMIMED). Cells are platedat 5×10⁴/well. After treatment of the cells, luciferase is measuredusing the Packard Luc Lite system (Cat. No.601691 1) according to themanufacturer's instructions in a final volume of 205 μl. Briefly, to acell suspension of 5×10⁵ cells/ml, γIFN (1000 U/ml Boehringer MannheimNo. 1050494) to a final concentration of 100 U/ml and 0.25% (v/v) LucLite Enhancer is added. After a 3 hour incubation LPS (50 μg/ml SIGMAL-8274) is added to give 5 μg/ml final concentration. The cells are thenplated at 5×10⁴/100 l/well into flat bottom white polystyrenemicroplates (Packard, Cat No. 6005180) and incubated for 16 hours. 5 μlof compound solution or control vehicle is then added and the cells arefurther incubated as indicated. 100 μl of luciferase substrate solutionis added and the plates are covered with TopSeal-A press-on adhesivesealing film (Packard Cat.No. 6005185) before measuring luminescencewith a Packard Top Count Scintillation Counter at 22° C. The luciferasesignal is stable for at least 90 min.

Example 2 Effect of the Radicicol Analog A

The THP-1 cell lines, clone No. 63 (containing PGL2_Neo30) and clone No.53 (containing pGL2-Control) are grown, differentiated and stimulatedwith γIFN and LPS identical to normal THP-1 cells. Radicicol analog A isadded 16 hours after the addition of LPS and cell extracts are thentaken 8 hours later or as indicated. Luciferase activity is inhibited by1 μM radicicol analog A on average by 50%±17%, in some cases inhibitionwas as great as 93%, whereas up to 5×10⁻⁶M of radical analog A has noeffects on the control clone No. 53, FIG. 3 (solid bars indicate cloneNo. 53, open bars clone No. 63).

Example 3 Application of Reporter Gene Assay to a Number of RadicicolAnalogs

A number of radicicol analogues are tested for their activity in thereporter assay substantially as described in the previous Examples. Theresults obtained are given in the Table below.

TABLE Luciferase reporter gene assay COMPOUND clone 0.5 μM 1 μM 5 μM

53 63 114  97 105  88 107  87

53 63  68  42  51  18  40  3

53 63  99  88  77  64  69  57

53 63  83  80  81  66  70  61

53 63 103 107 122  93 104  70

53 63 136  69 140  32 108  9

53 63  97  96  91  94  55  7

Example 4

Tablets, each comprising e.g. 50 mg of radicicol analog A or apharmaceutically acceptable salt, are prepared as follows:

Composition (10000 tablets) active ingredient 500.0 g lactose 500.0 gpotato starch 352.0 g gelatin  8.0 g talc  60.0 g magnesium stearate 10.0 g Silicon dioxide (highly dispersed)  20.0 g ethanol q.s.

The active ingredient is mixed with the lactose and 292 g of potatostarch and the mixture is moistened with an ethanolic solution of thegelatin and granulated through a sieve. After drying, the remainder ofthe potato starch, the magnesium stearate, the talc and the silicondioxide are mixed in and the mixture is compressed to form tablets, eachweighing 145.0 mg and comprising 50.0 mg of active ingredient; thetablets may, if desired, be provided with breaking notches for fineradaptation of the dose.

Example 5

Film-coated tablet, each comprising 100 mg of radicicol analog A or apharmaceutically acceptable salt are prepared as follows:

Composition (for 1000 film-coated tablets) active ingredient 100.0 g lactose 100.0 g  corn starch 70.0 g talc 60.0 g calcium stearate  1.5 ghydroxypropylmethylcellulose 2.36 g shellac 0.64 g water q.s  methylenechloride q.s.

The active ingredient, the lactose and 40 g of the corn starch are mixedand moistened with a paste prepared from 15 g of corn starch and water(with heating) and granulated. The granules are dried, the remainder ofthe corn starch, the talcum and the calcium stearate are added and mixedwith the granules. The mixture is compressed to form tablets (weight:280 mg) which are then film-coated with a solution of thehydroxypropylmethylcellulose and the shellac in methylene chloride;final weight of the film-coated tablet: 283 mg.

Example 6

Hard gelatin capsules, comprising 100 mg of active ingredient, forexample radicicol analog A or a pharmaceutically acceptable salt areprepared, for example, as follows:

Composition (for 1000 capsules) active ingredient 100.0 g lactose 250.0g microcrystalline cellulose  30.0 g sodium lauryl sulfate  2.0 gmagnesium stearate  8.0 g

The sodium lauryl sulfate is added to the lyophilised active ingredientthrough a sieve of 0.2 mm mesh size. The two components are intimatelymixed. Then first the lactose is added through a sieve of 0.6 mm meshsize and then the microcrystalline cellulose is added through a sieve of0.9 mm mesh size. The mixture is then intimately mixed again for 10minutes. Finally the magnesium stearate is added through a sieve of 0.8mm mesh size. After mixing for a further 3 minutes, size 0 hard gelatincapsules are each filled with 390 mg of the resulting formulation. Softgelatin capsules may be prepared using similar ingredients andprocedures.

What is claimed is:
 1. A method for the prophylaxis or treatment of adisease or medical condition having an etiology associated with theincreased stability of mRNA which contains one or more mRNA instabilitysequences, comprising administering to a human or animal patient aneffective amount of a compound of Formula II:

wherein R₁ is H, OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ loweralkyl-COO—; R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; R₃is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; —a—b— is—CHR₇—CHR₈— or cis or trans —CR₇═CR₈—; wherein R₇ and R₈ are the same ordifferent and are H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—,or —a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge; c is >CH—OH, >C═O or >CH₂; —d—e— is —CHR₇—CHR₈— or cis or trans—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and —f—g— is —CH₂—CH₂—,or cis or trans —CH═CH—, or a pharmaceutically acceptable salt thereofor a physiologically-hydrolysable and -acceptable ester thereof, whereinsaid compound induces degradation of the mRNA, provided that the diseaseor medical condition is not one with an etiology associated with orcomprising excessive cytokine release.
 2. A method for inducingdegradation of mRNA in a human or animal patient, which comprisesadministering to said patient an effective amount of a compound ofFormula II:

wherein R₁ is H, OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ loweralkyl-COO—; R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; R₃is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; —a—b— is-CHR₇—CHR₈— or cis or trans —CR₇═CR₈—; wherein R₇ and R₈ are the same ordifferent and are H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—,or —a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge; c is >CH—OH, >C═O or >CH₂; —d—e— is —CHR₇—CHR₈— or cis or trans—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and —f—g— is —CH₂—CH₂—,or cis or trans —CH═CH—, or a pharmaceutically acceptable salt thereofor a physiologically-hydrolysable and -acceptable ester thereof, whereinsaid compound induces mRNA degradation in the patient, and wherein themRNA contains an mRNA instability sequence, provided that the mRNA isnot mRNA coding for IL-1β, IL-6 or TNF-α.
 3. A method for theprophylaxis or treatment of a cancer or malignant disease in a human oranimal patient comprising administering to the patient an effectiveamount of a compound of Formula II:

wherein R₁ is OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ loweralkyl-COO—; R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; R₃is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; —a—b— is—CHR₇—CHR₈— or cis or trans —CR₇═CR₈—; wherein R₇ and R₈ are the same ordifferent and are H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—,or —a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge; c is >CH—OH, >C═O or >CH₂; —d—e— is —CHR₇—CHR₈— or cis or trans—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and —f—g— is —CH₂—CH₂—,or cis or trans —CH═CH—, or a pharmaceutically acceptable salt thereofor a physiologically-hydrolysable and -acceptable ester thereof.
 4. Amethod for the treatment or prevention of oncogene mediated cancers ormalignant diseases in a human or animal patient comprising administeringto the patient an effective amount of a compound of Formula II:

wherein R¹ is OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ loweralkyl-COO—; R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; R₃is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; —a—b— is—CHR₇—CHR₈— or cis or trans —CR₇=CR₈—; wherein R₇ and R₈ are the same ordifferent and are H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—,or —a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge; c is >CH—OH, >C═O or >CH₂; —d—e— is —CHR₇—CHR₈ — or cis or trans—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and —f—g— is —CH₂—CH₂—,or cis or trans —CH═CH—, or a pharmaceutically acceptable salt thereofor a physiologically-hydrolysable and -acceptable ester thereof.
 5. Amethod for the treatment or prevention of tumor growth or metastasis ina human or animal patient comprising administering to the patient aneffective amount of a compound of Formula II:

wherein R₁ is OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ loweralkyl-COO—; R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; R₃is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; —a—b— is—CHR₇—CHR₈— or cis or trans —CR₇═CR₈—; wherein R₇ and R₈ are the same ordifferent and are H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—,or —a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge; c is >CH—OH, >C═O or >CH₂; —d—e— is —CHR₇—CHR₈— or cis or trans—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and —f—g— is —CH₂—CH₂—,or cis or trans —CH═CH—, or a pharmaceutically acceptable salt thereofor a physiologically-hydrolysable and -acceptable ester thereof.
 6. Amethod to prevent or reverse multidrug resistance in a tumor cell in ahuman or animal patient comprising administering to the patient aneffective amount of a compound of Formula II:

wherein R₁ is OH, halogen, C₁-C₄ lower alkoxy, or C₁-C₄ loweralkyl-COO—; R₂ is OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; R₃is H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—; —a—b— is—CHR₇—CHR₈— or cis or trans —CR₇═CR₈—; wherein R₇ and R₈ are the same ordifferent and are H, OH, C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—,or —a—b— is —CHR₇—CHR₈— and R₇ and R₈ together with O form an epoxidebridge; c is >CH—OH, >C═O or >CH₂; —d—e— is —CHR₇—CHR₈— or cis or trans—CR₇═CR₈—, wherein R₇ and R₈ are the same or different and are H, OH,C₁-C₄ lower alkoxy, or C₁-C₄ lower alkyl-COO—, and —f—g— is —CH₂—CH₂—,or cis or trans —CH═CH—, or a pharmaceutically acceptable salt thereofor a physiologically-hydrolysable and -acceptable ester thereof.
 7. Themethod according to claim 3 or 4, wherein said cancer is breast cancer,colon cancer or lung cancer.